Team:NTHU Taiwan/Proof Of Concept
Introduction
Except for our experiment outcome, there are three components vital in our project. Firstly, we have to know whether selected genes are viable to kill pine wilt nematodes through RNAi. Secondly, which Agrobacterium strain is able to infect Pinus armandii. Last but not least, whether plants can express dsRNA is the most important aspect in the project. We had done paper studies to certify the above.
RNA interruption
We chose three different pine wilt nematodes’ genes. All of them are proven to kill nematodes through RNAi soaking. Two of the studies said that pine trees with RNAi soaking are more resistant to pine wilt nematodes compared to control groups. As a result, we realized that there are channels which can transport double strain RNA(dsRNA) among cells and lead to RNAi in nematodes.
| Genes | Function | Reference |
|---|---|---|
| BxPel | Pectate lyase | Xiu-Wen Qiu, Xiao-Qin Wu, Lin Huang, and Jian-Ren Ye. (2016) Influence of Bxpel1 Gene Silencing by dsRNA Interference on the Development and Pathogenicity of the Pine Wood Nematode, Bursaphelenchus xylophilus. Int J Mol Sci.17(1): 125. |
| BxPrx | Peroxiredoxin | Han-Yu Fu, Jia-Hong Ren, Lin Huang, Hao Li, and Jian-Ren Ye. (2014) Screening and Functional Analysis of the Peroxiredoxin Specifically Expressed in Bursaphelenchus xylophilus—The Causative Agent of Pine Wilt Disease. Int J Mol Sci,15(6): 10215–10232. |
| BxGPCR17454 | G protein-coupled receptor | Bowen Wang, Xin Hao, Jiayao Xu, Yan Ma, and Ling Ma1. (2019) Transcriptome-Based Analysis Reveals a Crucial Role of BxGPCR17454 in Low Temperature Response of Pine Wood Nematode. Int J Mol Sci, 20(12): 2898. |
Agrobacterium transformation
There are numerous commercial Agrobacterium strains in the world. However, not every strain is eligible to use in our project. We have to find the optimal strains to transform pine trees. Some papers reveal that the LBA4404 strain is able to infect Pinus armandii.
dsRNA expression in plants
There are few components for the RNAi mechanism, such as dicer, RISC complex and so on. Even though plants don’t hold the exactly same proteins, they have similar proteins to conduct the RNAi mechanism. Furthermore, there are several examples that plants express dsRNA to resist pathogens, like viruses or nematodes.
Conclusion
Our project aims to generate novel pine trees which are immune to pine wilt nematodes. Utilizing the RNAi mechanism as a method, we expected that pine expressed dsRNA to knot down nematodes’ genes. To achieve this goal, we had to understand detailed steps. Fortunately, these processes had been proven by some thesis. With these studies as backup, we believed that our innovation is not just an imagination but a feasible solution.
Reference
[1] Liu, X. Z.; Liu, Z.; Yang, Y. M.; Zhang, H. Y. Production of Transgenic Pinus Armandii Plants Harbouring BtCryIII(A) Gene. Biol. Plant. 2010, 54 (4), 711–714.
[2] Morozov, S. Y.; Solovyev, A. G.; Kalinina, N. O.; Taliansky, M. E. Double-Stranded RNAs in Plant Protection against Pathogenic Organisms and Viruses in Agriculture. Acta Naturae 2019, 11 (4), 13–21.
[3] Baulcombe, D. RNA Silencing in Plants. Nature 2004, 431 (7006), 356–363.
[4] Dutta, T. K.; Banakar, P.; Rao, U. The Status of RNAi-Based Transgenic Research in Plant Nematology. Front. Microbiol. 2014, 5, 760.